BCR or biomarker?

From the Warren laboratory, Clinical Research Division and Vaccine and Infectious Disease Division

Burkitt lymphoma (BL) is cancer of immune cells, specifically B cells. This cancer is the most common pediatric cancer in equatorial Africa. Tumors are fast growing and survival ranges from 30-50%. BL specifically affects B cells in the germinal center (antigen experienced B cells) and these cells usually show high levels of somatic hypermutation (SHM). SHM is the body’s natural way of confronting new antigens. It’s a process of controlled mutations in the region of the B cell receptor (BCR) that contacts the antigen, known as the variable region. BL also tends to involve B cells that have the IgM and IgD isotype BCR. In addition to SHM, B cells undergo immunoglobulin (Ig) rearrangement as part of their development, which results in a highly diverse repertoire. This process selects the “building blocks” of B cells and combines them at random. In order to identify which building blocks make up a certain B cell, sequencing is performed. Researchers believe that better understanding of the BCRs involved in BL could lead to better treatment and prevention strategies. Current work has focused on sequencing of tumor RNA which gives limited results based on productive and expressed Igs. In a recent study published in Blood Advances from Fred Hutchinson Clinical research and Vaccine and Infectious Disease Divisions, researchers used high-throughput sequencing (HTS) of tumor genomic DNA to study the complete Ig rearrangement in BL tumors.

Clonal relatedness score for BL and control samples. Overall, BL samples were more related compared to controls.
Clonal relatedness score for BL and control samples. Overall, BL samples were more related compared to controls.

In the current study, the authors sampled Ugandan and Ghanaian cohorts comprised of 51 BL patients. Tumor samples were subjected to HTS on genomic DNA to investigate the IGH, IGL, and IGK repertoire and rearrangements. The protocol used could capture unproductive, incomplete and complete rearrangements. Information generated from this procedure enabled the group to find a clonal rearrangement that comprised about 15% of the repertoire, compared to 2.25% in control reads (B cells from healthy PBMC and bone marrow). When looking at overall polyclonal nature of the populations, there was very little polyclonality in the BL population. At least one clonal locus was identified in 90% of BL tumors. When the sequences were compared by phylogenetic tree analysis, most of the unique sequences in each tumor were clustered in the tree and closely related to the tumor’s dominant sequence. To better define this, edit distance, which is the number of nucleotide differences between the tumor unique sequences and the most frequent sequence, was calculated. Edit distance calculations showed that BL tumor sequences were related with low-edit distance scores of less than 10 nucleotides which is low compared to controls. In addition to distance, clonal relatedness was calculated for each tumor (see figure). These data are defined as the fraction of unique sequences with less than a 10 edit distance from the most frequent sequences, resulting in most tumors having high relatedness. Comparing these data to the T cell receptor beta locus showed that this phenomenon is restricted to the Ig loci in tumor cells. This implies that mutations are not regulated by antigen-driven SHM. On a larger scale, the group also looked at allele usage and found five preferentially expressed alleles in BL BCRs as compared to healthy B cell controls. This suggests that BL progenitors with certain Ig genes may preferentially differentiate into the memory compartment. This study also suggests that these sequences may act as biomarkers to diagnose or monitor disease using the HTS protocol.

Lombardo KA, Coffey DG, Morales AJ, Carlson CS, Towlerton AMH, Gerdts SE, Nkrumah FK, Neequaye J, Biggar RJ, Orem J, Casper C, Mbulaiteye SM, Bhatia KG, Warren EH. 2017. High-throughput sequencing of the B-cell receptor in African Burkitt lymphoma reveals clues to pathogenesis. Blood Adv, 1(9), 535-544.

This work was funded through the National institutes of Health, National Cancer institute.

Research reported in the publication is a collaboration between Cancer Consortium members Christopher Carlson, Corey Casper, and Edus Warren.